There is an analysis technique that utilizes the so-called gradient elution method (a gradient elution technique) in which the composition (the constituent ratio) of an elute is sequentially changed and then flowed into a separation column using a high-performance liquid chromatograph (hereinafter, referred to as “HPLC”) to change the elution strength of a solute kept in the separation column.
This analysis technique has been widely applied because of its advantages in that a separation requiring a great elution time can be completed within a short time, a separation between molecules having similar structures can be allowed, and so on.
In this analysis technique, currently, it is general to feed the eluent at a flow velocity of about 1 ml/min. However, against the background of technical trends for increasing a sample-detecting accuracy by lowering the flow rate of a eluent in recent years, a HPLC being devised to feed the eluent at a flow velocity of as low as about 5 μl/min has been proposed.
However, the HPLC designed for low flow rate had a very complicated device structure, so that there was a basic technical problem in that the operation of the HPLC was difficult.
Furthermore, when a gradient elution is carried out under the conditions in which the flow velocity of the eluent is lower than 5 μl/min, it is possible to adapt a technology by which a flow path through which the eluent is transferred is split into a main stream and a subsidiary stream having a smaller inner diameter. Then, an analysis column is connected on the subsidary stream side, and then a gradient elution analysis is performed using an eluent flowing through the subsidary stream at a low flow velocity. However, this technology had technical problems in that a gradient elution can be hardly performed at high precision because the subsidary stream tended to be clogged, the desired low flow rate condition of the predetermined (Sub-Ul/min) was hardly kept in stable, and it became difficult to confirm whether the flowing was occurred at a predetermined flow rate.
Therefore, the present invention intends to provide a liquid chromatograph, an analysis system, and a method for gradient elution, which are designed so as to be allowed to make a change in gradient of an eluent at higher precision and to detect or analyze at high sensitivity by subsequently feeding the eluent under stable conditions of low flow velocity.